PROJECT SUMMARY Bacterial meningitis is the most common serious infection of the central nervous system (CNS) and a major cause of death and disability worldwide, especially in children. Although antibiotic therapy has changed bacterial meningitis from a uniformly fatal disease to an often curable one, the overall outcome remains unfavorable, with mortality of 5 to 10% and permanent neurologic sequelae occurring in 5 to 40% of survivors, depending on patient age and pathogen. Disruption and dysfunction of the blood-brain barrier (BBB) is a hallmark event in the pathophysiology of bacterial meningitis. Little is known, however, about the very first and crucial interaction between a bacterial pathogen with the BBB that initiates this chain of events, and may ultimately determine a poor or favorable neurological outcome in meningitis patients. This proposal seeks to elucidate the molecular mechanisms of BBB disruption during bacterial infection, and why it fails as a neuroprotective barrier during bacterial meningitis. We have shown that bacterial infection induces an epithelial to mesenchymal transition (EMT) program in endothelial cells (EndoMT), disrupting tight junctions in BBB endothelium through the upregulation of host transcription factor Snail1, a global repressor of tight junctions. Further, we have discovered that a bacterial adhesin interacts directly with vimentin, an intermediate filament protein that is induced during EMT/EndoMT. I hypothesize that BBB disruption may be due to the combined effect of bacterial entry and modulation of host signaling pathways that results in compromised barrier function. Further that bacterial pathogens associated with CNS disease possess the unique ability to penetrate brain endothelium, which ultimately leads to BBB dysfunction. These hypotheses will be addressed with both in vitro and in vivo models of BBB penetration using Group B streptococcus (GBS) as a model human pathogen associated with meningitis. AIM 1: Characterize the bacterial determinant(s) that initiate Snail1 activation and the contribution of Snail1 to BBB breakdown during GBS meningitis; AIM 2: Characterize the host factors that contribute to Snail1 activation during GBS infection; AIM 3: Determine the contribution of GBS-vimentin interaction to BBB penetration and the development of meningitis. These studies should increase our understanding of the bacterial and host factors involved in the interaction with brain endothelium that leads to barrier disruption, pathogen transit into the brain, and disease progression.